Euphorbia nigra extract and application of active ingredients thereof in preparation of medicine for treating chronic kidney disease

Abstract

The invention provides an application of an euphorbia nigra extract and an active component thereof in preparing a medicine for treating chronic nephropathy, which is characterized in that an adriamycin-induced chronic nephropathy rat model and a lipopolysaccharide (LPS), hydrogen peroxide (H2O2) and transforming growth factor-beta1 (TGF-beta1) induced HK-2 cell model are used as evaluation means; materials capable of remarkably treating kidney inflammation, oxidative stress and fibrosis are separated and identified from a chronic kidney disease treatment extract of beetle. From the aspects of regulating and controlling inflammation homeostasis, oxidative stress and fibrosis, inhibiting PI3K-Akt, MAPK and HIF-1 signal channels and the like, the effect and the action mechanism of the diving beetle extract and the 6-(3, 4-dihydroxyphenyl)-2, 3-naphthalenediol on treating the chronic kidney disease are elaborated, and the diving beetle extract and the 6-(3, 4-dihydroxyphenyl)-2, 3-naphthalenediol can be used as a chronic kidney disease treatment medicine and a related signal channel regulator.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomedicine, and particularly relates to the application of a Dysoptera exigua extract and active ingredients thereof in the preparation of a drug for treating chronic kidney disease. Background Art

[0002] Chronic kidney disease (CKD) is a serious health condition with a high prevalence, affecting approximately 10% of the global population. Fibrosis, oxidative stress, and inflammation are risk factors driving the development and progression of CKD. The PI3K-AKT, HIF-1, and MAPK signaling pathways are closely associated with fibrosis, oxidative stress, and inflammation. Inflammatory stimulation of the MAPK signaling pathway (including ERK and p38 kinase) can induce apoptosis in renal proximal tubular epithelial cells, promoting inflammation and renal damage. Oxidative stress is a key mechanism driving CKD progression, and the HIF-1 signaling pathway can regulate oxidative stress, leading to kidney damage. The PI3K-AKT signaling pathway is closely associated with the development and progression of renal fibrosis. Therefore, the development of clinical drugs and strategies to inhibit the PI3K-AKT, HIF-1, and MAPK signaling pathways for the treatment of CKD is essential.

[0003] Cybister chinensis Motschulsky is mentioned in numerous authoritative Chinese medical texts, including the Supplement to Compendium of Materia Medica. Extracts from Cybister chinensis Motschulsky have been shown to strengthen essence, enhance yang, nourish yin, tonify the kidneys, reduce urination, and activate blood circulation and remove blood stasis. However, there are currently no records of its use in the treatment of chronic kidney disease. Summary of the Invention

[0004] In view of this, the object of the present invention is to provide the use of a Chinese water slug extract and its active ingredients in the preparation of a drug for treating chronic kidney disease. The Chinese water slug extract and its active ingredients have a good inhibitory effect on chronic kidney disease inflammation, oxidative stress and fibrosis, and can be used as a drug for the treatment of chronic kidney disease and an inhibitor of PI3K-AKT, HIF-1 and MAPK signaling pathways.

[0005] The first aspect of the present invention aims to provide a use of a naphthalene diol derivative or a pharmaceutically acceptable salt thereof in the preparation of a drug for treating chronic kidney disease.

[0006] The second aspect of the present invention aims to provide a use of a Dytiscus euphorbia extract in the preparation of a drug for treating chronic kidney disease.

[0007] The third aspect of the present invention aims to provide a drug for treating chronic kidney disease.

[0008] The fourth aspect of the present invention aims to provide an inhibitor for chronic kidney disease-related signaling pathways.

[0009] To achieve the above purpose, the present invention is implemented through the following technical solutions

[0010] In a first aspect of the present invention, the present invention provides a use of a naphthalene diol derivative or a pharmaceutically acceptable salt thereof in the preparation of a drug for treating chronic kidney disease, wherein the naphthalene diol derivative is as shown in Formula I:

[0011]

[0012] in,

[0013] R1, R2, R 11 、R 12 Each is independently selected from hydrogen, C1-6 alkyl, C3-10 cycloalkyl, C1-6 acyl, phenyl or saccharyl;

[0014] R3, R4, R5, R6, R7, R8, R9, R 10 Each is independently selected from hydrogen, C1-6 alkyl, C3-10 cycloalkyl, C1-6 acyl, phenyl, saccharide, halogen, amino, hydroxyl, aldehyde or carboxyl.

[0015] In some embodiments of the present invention, the naphthalene diol derivative is 6-(3,4-dihydroxyphenyl)-2,3-naphthalene diol (Formula II).

[0016]

[0017] In a second aspect of the present invention, the present invention provides a use of a Dytiscus spp. extract in the preparation of a drug for treating chronic kidney disease, wherein the Dytiscus spp. extract comprises a naphthalene diol derivative described in Formula I or Formula II.

[0018] In some embodiments of the present invention, the chronic kidney disease includes chronic kidney disease induced by factors such as drugs, diabetes, hypertension, glomerulonephritis, etc.

[0019] In some embodiments of the present invention, the drug comprises doxorubicin, which can induce chronic kidney disease.

[0020] In some embodiments of the present invention, the chronic kidney disease includes chronic kidney disease caused by abnormal regulation of HK-2 in renal tubular epithelial cells induced by lipopolysaccharide (LPS), hydrogen peroxide (H2O2) or transforming growth factor-β1 (TGF-β1).

[0021] In some embodiments of the present invention, the chronic kidney disease includes chronic kidney disease caused by lipopolysaccharide (LPS)-induced abnormal regulation of HK-2 in renal tubular epithelial cells.

[0022] In some embodiments of the present invention, the chronic kidney disease includes chronic kidney disease caused by hydrogen peroxide (H2O2)-induced abnormal regulation of HK-2 in renal tubular epithelial cells.

[0023] In some embodiments of the present invention, the chronic kidney disease includes chronic kidney disease caused by abnormal regulation of HK-2 in renal tubular epithelial cells induced by transforming growth factor-β1 (TGF-β1).

[0024] In a third aspect, the present invention provides a drug for treating chronic kidney disease, wherein the active ingredients of the drug include a Dytiscus spp. extract and / or a naphthalene diol derivative.

[0025] In some embodiments of the present invention, the active ingredient of the medicament includes a Dytiscus edulis extract.

[0026] In some embodiments of the present invention, the active ingredient of the drug includes a naphthalene diol derivative.

[0027] In some embodiments of the present invention, the active ingredients of the drug include Dytiscus edulis extract and naphthalene diol derivatives.

[0028] In some embodiments of the present invention, the concentration of the naphthalene diol derivative in the in vitro cell model is 1 to 50 μM; in some embodiments of the present invention, the concentration of the naphthalene diol derivative in the in vitro cell model is 2.5 to 40 μM.

[0029] In some specific embodiments of the present invention, the concentration of the naphthalene diol derivative in the in vitro cell model is 1 μM; in some specific embodiments of the present invention, the concentration of the naphthalene diol derivative in the in vitro cell model is 2.5 μM; in some specific embodiments of the present invention, the concentration of the naphthalene diol derivative in the in vitro cell model is 5 μM; in some specific embodiments of the present invention, the concentration of the naphthalene diol derivative in the in vitro cell model is 10 μM; in some specific embodiments of the present invention, the concentration of the naphthalene diol derivative in the in vitro cell model is 15 μM; in some specific embodiments of the present invention, the concentration of the naphthalene diol derivative in the in vitro cell model is 20 μM; In some specific embodiments of the present invention, the concentration of the naphthalene diol derivative in the in vitro cell model is 25 μM; in some specific embodiments of the present invention, the concentration of the naphthalene diol derivative in the in vitro cell model is 30 μM; in some specific embodiments of the present invention, the concentration of the naphthalene diol derivative in the in vitro cell model is 35 μM; in some specific embodiments of the present invention, the concentration of the naphthalene diol derivative in the in vitro cell model is 40 μM; in some specific embodiments of the present invention, the concentration of the naphthalene diol derivative in the in vitro cell model is 45 μM; in some specific embodiments of the present invention, the concentration of the naphthalene diol derivative in the in vitro cell model is 50 μM.

[0030] In some embodiments of the present invention, the dosage of the Dytiscus edulis extract in animals is 50 to 300 mg / kg. In some embodiments of the present invention, the dosage of the Dytiscus edulis extract in animals is 55 to 220 mg / kg.

[0031] In some specific embodiments of the present invention, the dosage of the true water beetle extract in animals is 50 mg / kg; in some specific embodiments of the present invention, the dosage of the true water beetle extract in animals is 55 mg / kg; in some specific embodiments of the present invention, the dosage of the true water beetle extract in animals is 75 mg / kg; in some specific embodiments of the present invention, the dosage of the true water beetle extract in animals is 100 mg / kg; in some specific embodiments of the present invention, the dosage of the true water beetle extract in animals is 110 mg / kg; in some specific embodiments of the present invention, the dosage of the true water beetle extract in animals is 125 mg / kg; In some specific embodiments of the present invention, the dosage of the true water beetle extract in animals is 150 mg / kg; in some specific embodiments of the present invention, the dosage of the true water beetle extract in animals is 175 mg / kg; in some specific embodiments of the present invention, the dosage of the true water beetle extract in animals is 200 mg / kg; in some specific embodiments of the present invention, the dosage of the true water beetle extract in animals is 220 mg / kg; in some specific embodiments of the present invention, the dosage of the true water beetle extract in animals is 250 mg / kg; in some specific embodiments of the present invention, the dosage of the true water beetle extract in animals is 300 mg / kg.

[0032] In the fourth aspect of the present invention, the present invention provides an inhibitor for chronic kidney disease-related signal pathways, the active ingredients of the inhibitor include a true water beetle extract and / or a naphthalene diphenol derivative, and the signal pathways include phosphatidylinositol 3-kinase (PI3K)-protein kinase B (Akt), mitogen-activated protein kinase (MAPK), and hypoxia-inducible factor-1 (HIF-1) signal pathways.

[0033] In some embodiments of the present invention, the concentration of the naphthalene diol derivative is 2.5 to 40 μM; the concentration of the Dysoptera exigua extract is 55 to 220 mg / kg.

[0034] Beneficial effects: The present invention provides the use of Chinese water beetle extracts and naphthalene diol derivatives in the preparation of drugs for the treatment of chronic kidney disease. The present invention uses the chronic kidney disease rat model induced by doxorubicin and the HK-2 cell model induced by lipopolysaccharide (LPS), hydrogen peroxide (H2O2), and transforming growth factor-β1 (TGF-β1) as evaluation methods, and isolates and identifies a monomer substance with significant therapeutic effects on renal inflammation, oxidative stress and fibrosis from the water beetle extract for treating chronic kidney disease: 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol, i.e. 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol. The evaluation results show that the water beetle extract can improve rat kidney pathological damage, regulate the levels of inflammatory factors, oxidative stress and fibrosis indicators. 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol can effectively inhibit inflammation, oxidative stress and fibrosis-related indicators. The present invention describes the effects and mechanisms of Dysphagnum pectinum extract and 6-(3,4-dihydroxyphenyl)-2,3-naphthalene diol in treating chronic kidney disease from the aspects of regulating inflammatory homeostasis, oxidative stress and fibrosis, and inhibiting PI3K-Akt, MAPK and HIF-1 signaling pathways. The Dysphagnum pectinum extract and 6-(3,4-dihydroxyphenyl)-2,3-naphthalene diol can be used as drugs for treating chronic kidney disease and regulators of related signaling pathways. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Figure 1 This figure shows the effect of Dysthisis extract on urine protein content in CKD rats (the low, medium and high dose groups of Dysthisis extract are referred to as L, M and H, respectively, the same below);

[0036] Figure 2 This is the effect of Dysthisis extract on serum protein levels in CKD rats;

[0037] Figure 3 Effects of Dysthisis extract on serum urea nitrogen (BUN) and creatinine (Scr) levels in CKD rats:

[0038] Figure 4 This is a graph showing the effects of Dysthisis extract on blood lipid indicators in CKD rats;

[0039] Figure 5 This is the effect of Dysthisis extract on IL-6 and IL-1β in CKD rats;

[0040] Figure 6 This is a graph showing the effect of Dysthisis extract on hematoxylin-eosin (HE) and Masson staining of kidney tissue in CKD rats;

[0041] Figure 7 The effect of Dysthisis extract on ROS fluorescence staining in kidney tissue of CKD rats

[0042] Figure 8This figure shows the effect of Dysthisis extract on immunohistochemical staining of α-SMA and E-cadherin in kidney tissue of CKD rats;

[0043] Figure 9 This is a diagram showing the effect of Dysthisis extract on the levels of proteins related to the PI3K-AKT signaling pathway;

[0044] Figure 10 This is a diagram showing the effect of Dysthisis extract on the levels of proteins related to the MAPK signaling pathway;

[0045] Figure 11 This is a diagram showing the effect of Dysthisis extract on the levels of proteins related to the HIF-1 signaling pathway;

[0046] Figure 12 This is a graph showing the effect of 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol (referred to as compound 1 in the figure, the same below) on LPS-induced TNF-α, IL-6, and IL-1β factors in HK-2 cells;

[0047] Figure 13 Graph showing the effect of 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol on the p-ERK protein in HK-2 cells induced by LPS;

[0048] Figure 14 The figure shows the effect of 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol on GSH, SOD and MDA of HK-2 cells induced by H2O2;

[0049] Figure 15 Graph showing the effect of 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol on ROS in HK-2 cells induced by H2O2;

[0050] Figure 16 The figure shows the effect of 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol on HIF-1α protein in HK-2 cells induced by H2O2;

[0051] Figure 17 The figure shows the effect of 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol on α-SMA, COL-Ⅰ and E-cadherin in HK-2 cells induced by TGF-β1;

[0052] Figure 18This figure shows the effect of 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol on TGF-β1-induced p-PI3K and P-AKT proteins in HK-2 cells.

[0053] Specific implementation (embodiment)

[0054] In order to enable those skilled in the art to better understand the present invention, the following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments in the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts should fall within the scope of protection of this application.

[0055] It should be noted that the terms "including" and "having" and any variations thereof in the specification and claims of this application and the above-mentioned drawings are intended to cover non-exclusive inclusions. For example, a process, method, system, product or apparatus that includes a series of steps or units is not necessarily limited to those steps or units clearly listed, but may include other steps or units that are not clearly listed or are inherent to these processes, methods, products or apparatuses.

[0056] The present invention does not specifically limit the source of the Dytiscus chinensis extract, and it is preferably derived from the School of Pharmacy of Dali University. The naphthalene diphenol derivatives shown in the present invention are preferably extracted from the Dytiscus chinensis extract, and the extraction method preferably comprises: (1) using the extract of the Dytiscus chinensis extract to obtain 17 components through silica gel normal phase column chromatography (petroleum ether: ethyl acetate, 100:0 to 0:100); (2) wherein component Fr.4 is subjected to silica gel normal phase column chromatography (chloroform: methanol, 100:0 to 0:100) to obtain component Fr.4.7; (3) component Fr.4.7 is subjected to LH-20 gel column chromatography (chloroform: methanol, 1:1) to obtain the target product naphthalene diphenol derivatives.

[0057] In an embodiment of the present invention, the effect of the Dysopus extract was verified by constructing a rat model of chronic kidney disease induced by doxorubicin. The method for constructing the rat model of chronic kidney disease induced by doxorubicin preferably comprises the following steps: selecting healthy male SD rats, acclimating them to feeding for one week, and then injecting doxorubicin into the tail vein at a dose of 4 mg / kg. One week later, the rats were injected with doxorubicin into the tail vein at a dose of 3 mg / kg to establish the chronic kidney disease model.

[0058] When the present invention utilizes the doxorubicin-induced chronic kidney disease rat model to verify its function, it is preferred that the successfully modeled chronic kidney disease rats be treated with the Dysoptera exigua extract and corresponding controls, and the effects are evaluated by detecting relevant indicators. The markers described in the present invention include: indicators of renal tissue pathological changes, including changes in glomerular and tubular structure and fibrillar collagen deposition; renal function indicators, including urine protein, serum protein (ALB), serum urea nitrogen (BUN) and serum creatinine (Scr); inflammation-related indicators, including serum interleukin-6 (IL-6), interleukin-1β (IL-1β), phosphorylated extracellular signal-regulated kinase (p-ERK), phosphorylated p38 mitogen-activated protein kinase (p-p38) and phosphorylated c-Jun amino-terminal kinase (p-JNK); oxidative stress indicators, including reactive oxygen species (ROS) and HIF-1α; lipid metabolism indicators, including serum total cholesterol (TC) and triglycerides (TG); and fibrosis markers, including α-smooth muscle actin (α-SMA), epithelial cadherin (E-cadherin), p-PI3K, p-AKT and phosphorylated mammalian target of rapamycin (p-mTOR). The present invention preferably detects by hematoxylin-eosin (HE) staining, Masson staining, enzyme-linked immunosorbent assay (ELISA), immunofluorescence, immunohistochemistry and western blot.

[0059] In an embodiment of the present invention, the effects of the naphthalene diol derivatives of Dysphaeranthus were verified by constructing an HK-2 cell model induced by LPS, H2O2, and TGF-β1. The method for constructing the cell model preferably includes the following steps:

[0060] LPS-induced inflammatory cell model: HK-2 cells in the logarithmic growth phase were selected and stimulated with LPS at a final concentration of 40 μg / mL for 24 hours to induce an inflammatory cell model.

[0061] H2O2-induced oxidative stress cell model: HK-2 cells in the logarithmic growth phase were stimulated with 150μM H2O2 for 4h to construct an oxidative stress cell model.

[0062] TGF-β1-induced fibrosis cell model: HK-2 cells in the logarithmic growth phase were stimulated with TGF-β1 at a final concentration of 10 ng / mL for 24 h to establish a fibrosis cell model.

[0063] When the present invention utilizes the above-mentioned cell model to verify the function, it is preferred that the constructed cell model be co-incubated with different concentrations of naphthalene diol derivatives, and the effect be evaluated by detecting relevant markers. The markers described in the present invention include: inflammatory markers including tumor necrosis factor-α (TNF-α), IL-6, IL-1β, and p-ERK; oxidative stress markers including glutathione (GSH), superoxide dismutase (SOD), malondialdehyde (MDA), ROS, and HIF-1α; and fibrosis markers including α-SMA, type I collagen (COL-I), E-cadherin, p-PI3K, and p-AKT. The present invention preferably uses ELISA, immunofluorescence, real-time reverse transcription polymerase chain reaction (RT-PCR), flow cytometry, and western blot for detection.

[0064] The following are some definitions used in this invention:

[0065] In the present invention, "Cybister extract" refers to a mixture obtained from adults or larvae of the genus Cybister, a member of the family Dytiscidae, by physical disruption, solvent extraction (e.g., ethanol, water, methanol), chromatographic separation, and the like, including extracts of Cybister chinensis (i.e., from species of Cybister distributed in China, such as Cybister chinensis Motschulsky). The "Cybister extract" described in the present invention specifically refers to extracts of Cybister chinensis (e.g., Cybister chinensis Motschulsky).

[0066] In the present invention, the "administered dose" refers to the amount of a drug administered per unit body weight of a mammal (preferably a rat or a human).

[0067] In the present invention, "concentration in an in vitro cell model" refers to the final mass concentration or molar concentration of the test substance (such as naphthalene diol derivatives, Dysoptera extract, etc.) in the cell culture medium in an in vitro cell culture system (such as adherent cells or suspension cell culture), and the unit is usually μg / mL, mg / mL or μmol / L. In the present invention, the concentration in the in vitro cell model adopts μM (micromoles / liter) as the standard unit.

[0068] In the present invention, "inhibitors of signaling pathways" refer to compounds or biological molecules that can directly or indirectly inhibit the activity of specific signal transduction pathways in cells, by blocking the expression, phosphorylation, interaction or enzymatic activity of key proteins in the pathway (such as receptors, kinases, transcription factors, etc.), thereby inhibiting the activation of the pathway.

[0069] In the present invention, "active ingredient" refers to a single chemical entity, biological molecule or mixture that can directly produce the expected pharmacological action, physiological activity or biological effect in a drug, health product, cosmetic or biological product. In the present invention, "active ingredient" refers to a substance with pharmacological activity for treating chronic kidney disease (CKD), including but not limited to: naphthalene diol derivatives: the structure is shown in general formula (I), preferably 6-(3,4-dihydroxyphenyl)-2,3-naphthalene diol, and its pharmaceutically acceptable salts, solvates or prodrug forms; true water beetle extract: refers to the biologically active component obtained by extraction from the Chinese true water beetle (Cybister chinensis) with a solvent (such as water, ethanol).

[0070] The present invention will be further analyzed below with reference to specific examples.

[0071] Example 1

[0072] Effects of Dytiscidae Extract on Urinary Protein Content in CKD Rats

[0073] Experimental method: 6 rats were randomly selected as the normal group, and the remaining rats were used for modeling. After modeling, the rats were randomly divided into the model group, the nephritis recovery tablets group (750 mg / kg), the high-dose group of water beetle extract (220 mg / kg), the medium-dose group of water beetle extract (110 mg / kg) and the low-dose group of water beetle extract (55 mg / kg). On the first day after grouping, the rats in each group were gavaged with the corresponding drugs. The normal control group and the model control group were given an equal volume of 0.5% sodium carboxymethyl cellulose and 1% Tween 80 solution prepared in 0.9% normal saline, once a day, for 4 consecutive weeks (subsequent Examples 2-11 all adopted this treatment).

[0074] During the 4 weeks of continuous administration, the rats were placed in clean metabolic cages at the end of each week and fasted but not watered. 24-hour urine was collected and the 24-hour urine protein content was determined by the Brandford method.

[0075] The experimental results are as follows Figure 1 As shown in the results: before modeling (week 0), there was no statistical difference in the 24-hour urine protein content of rats in each group, that is, the status of rats in each group was the same; 3 weeks after modeling, the 24-hour urine protein content of rats in each modeling group increased significantly and the 24-hour urine protein content was >50 mg·24h, that is, the CKD rat model was successfully established; after 4 weeks of drug treatment, the 24-hour urine protein content of rats in each treatment group was significantly reduced (p < 0.01 ~ 0.05), indicating that the water beetle extract has a good effect in reducing urine protein.

[0076] Example 2

[0077] Effects of Dysthisis Extract on Serum ALB Levels in CKD Rats

[0078] Experimental Methods: After the final dose, rats were fasted for 24 hours, anesthetized with isoflurane, and blood was collected from the abdominal aorta. After 4 hours of incubation at 4°C, the blood was centrifuged at 3000 rpm for 10 minutes. The upper serum layer was collected and aliquoted into EP tubes and stored at -80°C. Serum protein content was determined using a serum ALB kit.

[0079] The experimental results are as follows Figure 2 As shown in the results: compared with the normal control group, the serum ALB of rats in the model control group was significantly decreased (p<0.01); compared with the model control group, the serum ALB of rats in the low-dose water beetle extract group (p<0.05), the nephritis recovery tablets group, and the high- and medium-dose water beetle extract groups were all increased, indicating that water beetle extract has an improving effect on the filtration function of the kidneys of CKD rats.

[0080] Example 3

[0081] Effects of Dyschisis Extract on Serum Urea Nitrogen (BUN) and Serum Creatinine (Scr) in CKD Rats

[0082] Experimental Methods: After the final dose, rats were fasted for 24 hours, anesthetized with isoflurane, and blood was collected from the abdominal aorta. After 4 hours of incubation at 4°C, the blood was centrifuged at 3000 rpm for 10 minutes. The upper serum layer was collected and aliquoted into EP tubes and stored at -80°C. BUN and Scr levels were measured using BUN and Scr kits.

[0083] The experimental results are as follows Figure 3 As shown in the results: compared with the normal control group, the serum BUN and Scr of the rats in the model control group were significantly increased (p<0.01~0.05); compared with the model control group, the serum Scr of the rats in the Nephritis Recovery Tablets group and the Dysthisis extract groups were significantly decreased (p<0.01~0.05), and the serum BUN of the rats in the Nephritis Recovery Tablets group and the Dysthisis extract low (p<0.01), medium and high dose groups (p<0.01) were decreased to varying degrees, indicating that the Dysthisis extract can alleviate the renal function damage of CKD rats.

[0084] Example 4

[0085] Effects of Dytiscidae Extract on Blood Lipid Indexes in CKD Rats

[0086] Experimental Methods: Rats were fasted for 24 hours after the final dose, but not water. Blood was collected from the abdominal aorta and allowed to rest at 4°C for 4 hours. The blood was then centrifuged at 3000 rpm for 10 minutes. The upper serum layer was collected and aliquoted into EP tubes and stored at -80°C. Serum total cholesterol (TC) and triglyceride (TG) levels were measured using TC and TG kits.

[0087] The experimental results are as follows Figure 4As shown in the results: compared with the normal control group, the serum TC and TG contents of the rats in the model control group were significantly increased (p<0.0001); compared with the model control group, the serum TC of the rats in the Nephritis Rehabilitation Tablets group and the low (p<0.05), medium (p<0.05) and high dose groups of the Dysthisis extract were decreased, and the serum TG of the rats in the Nephritis Rehabilitation Tablets group and the low (p<0.01), medium (p<0.05) and high (p<0.05) dose groups of the Dysthisis extract were decreased. The results showed that the Dysthisis extract can effectively reduce the serum lipid levels of CKD rats and improve lipid metabolism disorders in CKD rats.

[0088] Example 5

[0089] Effects of Dyschisis Extract on IL-6 and IL-1β in CKD Rats

[0090] Experimental Methods: After the final dose, rats were fasted for 24 hours, anesthetized with isoflurane, and blood was collected from the abdominal aorta. After 4 hours of incubation at 4°C, the blood was centrifuged at 3000 rpm for 10 minutes. The upper serum layer was collected and aliquoted into EP tubes and stored at -80°C. IL-6 and IL-1β levels were determined using IL-6 and IL-1β kits.

[0091] The experimental results are as follows Figure 5 As shown in the results: Compared with the normal control group, the serum IL-6 and IL-1β levels of rats in the model control group were significantly increased (p<0.0001); compared with the model control group, the serum IL-6 and IL-1β levels of rats in the Nephritis Recovery Tablets group and the Dysthisis extract groups were decreased (p<0.0001). The results showed that Dysthisis extract can effectively reduce the serum inflammatory factor levels of CKD rats.

[0092] Example 6

[0093] Effects of Dyschisis Extract on Hematoxylin-Eosin (HE) and Masson's Staining of Kidney Tissue in CKD Rats

[0094] Experimental Methods: Kidneys were fixed with paraformaldehyde (4%), then dehydrated in ethanol and embedded in paraffin. 4-μm paraffin sections were prepared for H&E and Masson staining. The degree of renal injury was graded from 0 to 3 by examining tubular lesions, which included tubular atrophy and necrosis, inflammatory cell infiltration, and interstitial fibrosis. The scores for each criterion were as follows: 0 = none; 1 = mild (<25%); 2 = moderate (25%-50%); and 3 = severe (>50%). The degree of fibrosis was assessed by light microscopy based on the amount of collagen deposited in the kidneys.

[0095] The experimental results are as follows Figure 6As shown: HE staining results showed that the morphology and structure of the renal tubules and glomeruli of rats in the normal control group were normal, with occasional congestion in the glomeruli and occasional inflammatory cell infiltration in the renal interstitium; the morphology and structure of the renal tubules and glomeruli of rats in the model control group were destroyed, the renal tubular lumen was dilated, the glomeruli were often congested, and a large number of inflammatory cells infiltrated in the renal interstitium; the renal tubular lumen was slightly dilated in the Nephritis Recovery Tablet group and the water beetle extract treatment groups, with occasional congestion in the glomeruli and occasional inflammatory cell infiltration in the renal interstitium. Masson staining results showed that the area of collagen fibrosis in the renal tissue of rats in the model group was significantly increased, collagen staining in the glomeruli increased, and the basement membrane thickness increased, indicating that the model group rats had kidney damage and a high degree of renal fibrosis; while in the renal tissue of the Nephritis Recovery Tablet group and the water beetle extract treatment groups, the area of glomerular collagen fiber staining decreased and the basement membrane thickness improved, indicating that water beetle extract can delay renal fibrosis in CKD rats. Semi-quantitative pathological scoring revealed that the renal tubular injury score in the model group was significantly higher than that in the normal group (p<0.0001), while the renal tubular injury score in the group treated with Dysphid extract was significantly reduced (p<0.0001). Masson staining revealed a significant increase in the area of collagen deposition in the renal tissue of rats in the model group (p<0.0001), while the area of collagen deposition in the group treated with Dysphid extract was significantly reduced (p<0.0001). These results indicate that Dysphid extract can alleviate renal injury and renal fibrosis.

[0096] Example 7

[0097] Effects of Dyschisis Extract on ROS Fluorescence Staining in Kidney Tissue of CKD Rats

[0098] Experimental method: Kidneys were fixed with paraformaldehyde (4%), dehydrated in ethanol, and embedded in paraffin. 4-micron paraffin sections were prepared for ROS fluorescence staining.

[0099] The experimental results are as follows Figure 7 As shown in the results: Compared with the normal control group, the ROS level of rats in the model control group was significantly increased (p < 0.0001), while the Shenyan Kangfu Tablets and Dysthisis extract groups at all doses had a significant inhibitory effect on the oxidative damage caused by CKD (p < 0.001 ~ 0.01).

[0100] Example 8

[0101] Effects of Dyschisis Extract on Immunohistochemical Staining of α-SMA and E-cadherin in Kidney Tissue of CKD Rats

[0102] Experimental Methods: Kidneys were fixed with paraformaldehyde (4%), dehydrated in ethanol, and embedded in paraffin. 4-μm paraffin sections were prepared for immunohistochemical staining of α-smooth muscle actin (α-SMA) and epithelial cadherin (E-cadherin).

[0103] The experimental results are as follows Figure 8 As shown in the results of immunohistochemical staining, compared with the normal group, the expression of α-SMA in the model group was significantly upregulated, and the expression of E-cadherin was significantly downregulated (p < 0.0001); after treatment with D. diversifolia extract, the upregulation of α-SMA was reduced (p < 0.0001), and the downregulation of E-cadherin was reduced (p < 0.001 ~ 0.0001). These findings indicate that D. diversifolia extract has an anti-fibrotic effect on CKD rats.

[0104] Example 9

[0105] Effects of Dyschisis Extract on the Levels of Proteins Related to the Phosphatidylinositol 3-Kinase (PI3K-AKT) Signaling Pathway

[0106] Experimental method: Kidneys were fixed with paraformaldehyde (4%), dehydrated in ethanol, and embedded in paraffin. 4-μm paraffin sections were prepared for immunofluorescence staining of p-PI3K, p-AKT, and p-mTOR.

[0107] The experimental results are as follows Figure 9 As shown: Immunofluorescence assay results showed that compared with the normal group, the protein levels of p-PI3K, p-AKT, and p-mTOR in the renal tissue of rats in the model group were significantly increased (p<0.0001), indicating that the PI3K-Akt signaling pathway is activated in the CKD model. Compared with the model group, the protein levels of p-PI3K, p-AKT, and p-mTOR in the rats treated with Dysphid extract were significantly decreased (p<0.0001-0.01). This indicates that Dysphid extract can effectively inhibit the overactivation of the PI3K-Akt signaling pathway, thereby exerting its anti-fibrotic effects.

[0108] Example 10

[0109] Effects of Dysthisis Extract on the Levels of Proteins Related to MAPK Signaling Pathway

[0110] Experimental methods such as Figure 10 Shown: Kidneys were fixed with paraformaldehyde (4%), dehydrated in ethanol, and embedded in paraffin. 4-μm paraffin sections were prepared for immunofluorescence staining of phosphorylated extracellular signal-regulated kinase (p-ERK), phosphorylated c-Jun N-terminal kinase (p-p38), and phosphorylated p38 mitogen-activated protein kinase (p-JNK).

[0111] Experimental Results: Immunofluorescence assays showed that compared with the normal group, the protein levels of phosphorylated extracellular signal-regulated kinase (p-ERK), phosphorylated c-Jun N-terminal kinase (p-p38), and phosphorylated p38 mitogen-activated protein kinase (p-JNK) in the renal tissues of rats in the model group were significantly increased (p<0.0001), indicating that the MAPK signaling pathway is activated in the CKD model. Compared with the model group, the protein levels of p-ERK, p-p38, and p-JNK in the treatment groups treated with D. edulis extract were decreased to varying degrees (p<0.0001). This suggests that D. edulis extract can effectively inhibit the overactivation of the MAPK signaling pathway, thereby exerting an anti-inflammatory effect.

[0112] Example 11

[0113] Effects of Dysphaeria melongena extract on the levels of proteins related to the phosphorylation of hypoxia-inducible factor (HIF-1) signaling pathway

[0114] Experimental method: Kidneys were fixed with paraformaldehyde (4%), dehydrated in ethanol, and embedded in paraffin. 4-μm paraffin sections were prepared for HIF-1α immunofluorescence staining.

[0115] The experimental results are as follows Figure 11 The experimental results show that compared with the normal group, the HIF-1α protein level in the renal tissue of rats in the model group was significantly increased (p<0.0001), indicating that the HIF-1 signaling pathway is activated in the CKD model. Compared with the model group, the HIF-1α protein level in the rats treated with the water beetle extract was significantly decreased (p<0.0001). This indicates that the water beetle extract can effectively inhibit the overactivation of the HIF-1 signaling pathway, thereby exerting its antioxidant effect.

[0116] Example 12

[0117] Effects of 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol on LPS-induced TNF-α, IL-6, and IL-1β in HK-2 cells

[0118] Experimental method: HK-2 cells in the logarithmic growth phase were selected and digested with trypsin to prepare a concentration of 3×10 5A cell suspension of 100 μg / mL was seeded in 6-well plates. Experimental, negative, and blank groups were set up, and all plates were incubated overnight at 37°C in a 5% CO2 incubator. Experimental groups were stimulated with 40 μg / mL LPS for 24 hours. 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol was prepared in complete culture medium at concentrations of 2.5 μM, 5 μM, and 10 μM. An equal volume of culture medium was added to the control group. Each group was treated with triplicate wells for 24 hours. Cells were gently washed with cold PBS, then trypsinized and harvested by centrifugation at 1000 × g for 5 minutes. The harvested cells were washed three times with cold PBS, resuspended in 150 μL PBS, and disrupted by sonication. The extract was centrifuged at 1500 × g for 10 minutes at 2-8°C. The supernatant was collected and assayed for TNF-α, IL-6, and IL-1β levels according to the kit instructions.

[0119] The experimental results are as follows Figure 12 As shown: Compared with the control group, the levels of TNF-α, IL-6, and IL-1β in the model group were significantly increased (p<0.0001), indicating that LPS can induce inflammatory responses in cells. Compared with the model group, the levels of TNF-α, IL-6, and IL-1β were significantly decreased after treatment with 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol (p<0.0001). The experimental results show that 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol has a significant inhibitory effect on the LPS-induced inflammatory response in HK-2 cells, demonstrating significant anti-inflammatory activity.

[0120] Example 13

[0121] Effects of 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol on LPS-induced phosphorylation of extracellular signal-regulated kinase (p-ERK) in HK-2 cells

[0122] Experimental method: HK-2 cells in the logarithmic growth phase were selected and digested with trypsin to prepare a concentration of 3×10 5A cell suspension of 100 μg / mL was seeded in 6-well plates. Experimental, negative, and blank groups were set up, and all plates were incubated overnight at 37°C in a 5% CO2 incubator. Experimental groups were stimulated with 40 μg / mL LPS for 24 hours. 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol was prepared in complete culture medium at concentrations of 5 μM, 10 μM, and 20 μM. The control group received an equal volume of culture medium. Each group was treated with three replicates for 24 hours. The supernatant was discarded, and the culture medium was washed thoroughly with PBS. Pre-prepared cell lysis buffer was added, with 150 μL of lysis buffer added to each well. Mix thoroughly by pipetting, and then refrigerate at 4°C for half an hour to fully lyse the cells. After lysis, the cells were transferred to centrifuge tubes and centrifuged at 12,000 rpm for 10 minutes at 4°C. The supernatant was collected to obtain total protein. One-quarter volume of loading buffer was added, and the cells were boiled at 100°C for 5 minutes before aliquoting and storing at -80°C for later use. Protein concentration was measured using a BCA kit. After preparing the electrophoresis tape, add the marker and sample for electrophoresis. After electrophoresis, cut the gel containing the target protein and transfer it to a membrane. After transfer, wash the membrane with TBST and block it with blocking solution for 1 hour. Incubate with diluted primary antibody overnight. After washing with TBST, incubate with secondary antibody at room temperature for 1 hour. Wash the membrane again with TBST, develop with ECL developer, acquire images, and measure p-ERK levels.

[0123] The experimental results are as follows Figure 13 As shown: Compared with the control group, the expression of p-ERK protein in the model group increased; compared with the model group, the expression of p-ERK protein in the 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol-treated groups (5, 10 μM) decreased, and the expression of p-ERK protein was significantly reduced at a concentration of 20 μM (p<0.05), showing a dose-dependent manner.

[0124] Example 14

[0125] Effects of 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol on glutathione (GSH), superoxide dismutase (SOD) and malondialdehyde (MDA) in HK-2 cells induced by H2O2

[0126] Experimental method: HK-2 cells in the logarithmic growth phase were selected and digested with trypsin to prepare a concentration of 3×10 5A cell suspension of 100 μg / mL was seeded in 6-well plates. Experimental, negative, and blank groups were set up, and all plates were incubated overnight in a 37°C, 5% CO2 incubator. 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol was prepared in complete culture medium at concentrations of 5μM, 10μM, and 20μM. An equal volume of complete culture medium was added to the control group, with triplicate wells per group. Models were established after 4 hours of drug stimulation. Experimental groups were stimulated with 150μM H2O2 for 4 hours. GSH, SOD, and MDA levels in each group were analyzed according to the kit instructions.

[0127] The experimental results are as follows Figure 14 As shown in the results, compared with the control group, the GSH level in the model group was significantly decreased (p<0.0001), while the MDA and SOD levels were significantly increased (p<0.0001), indicating H2O2-induced oxidative stress. Treatment with 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol significantly increased GSH levels (p<0.001-0.01), while MDA and SOD levels were significantly decreased (p<0.0001-0.01). This indicates that 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol can effectively alleviate H2O2-induced oxidative damage and enhance the antioxidant capacity of cells. In summary, 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol has a protective effect against H2O2-induced damage in HK-2 cells.

[0128] Example 15

[0129] Effects of 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol on H2O2-induced reactive oxygen species (ROS) in HK-2 cells

[0130] Experimental method: HK-2 cells in the logarithmic growth phase were selected and digested with trypsin to prepare a concentration of 3×10 5 A cell suspension of 100 μg / mL was seeded in 6-well plates. Experimental, negative, and blank groups were set up, and all plates were incubated overnight in a 37°C, 5% CO2 incubator. 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol was added to complete culture medium at concentrations of 5 μM, 10 μM, and 20 μM. An equal volume of complete culture medium was added to the control group, with triplicate wells per group. Models were established 4 hours after drug stimulation. Immunofluorescence analysis was performed according to the kit instructions to analyze ROS levels in each cell group.

[0131] The experimental results are as follows Figure 15 As shown in the results: Compared with the control group, H2O2 (150μM) stimulation significantly increased the intracellular ROS level (p<0.0001). Compared with the model group, the cellular ROS level in the 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol-treated group was significantly reduced (p<0.0001), indicating that 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol can effectively inhibit the generation of ROS induced by H2O2 and has a certain protective effect against oxidative damage.

[0132] Example 16

[0133] Effects of 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol on H2O2-induced HIF-1α in HK-2 cells

[0134] Experimental method: HK-2 cells in the logarithmic growth phase were selected and digested with trypsin to prepare a concentration of 1×10 6 A cell suspension of 100 μL / mL was seeded in a 6-well plate. Experimental, negative, and blank groups were set up, and all plates were incubated overnight at 37°C in a 5% CO2 incubator. 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol was prepared in complete culture medium at concentrations of 5 μM, 10 μM, and 20 μM. An equal volume of complete culture medium was added to the control group, with triplicate wells per group. Models were established 4 hours after drug stimulation. The supernatant was discarded, and the culture medium was washed thoroughly with PBS. Pre-prepared cell lysis buffer was added, with 150 μL added to each well. Mix thoroughly by pipetting, and then refrigerate at 4°C for half an hour to fully lyse the cells. After lysis, the cells were transferred to a centrifuge tube and centrifuged at 12,000 rpm for 10 minutes at 4°C. The supernatant was collected to obtain total protein. One-quarter volume of loading buffer was added, and the cells were boiled at 100°C for 5 minutes before aliquoting and storing at -80°C. Protein concentration was determined using a BCA assay kit. Prepare the electrophoresis tape, add the marker and sample, and perform electrophoresis. After electrophoresis, cut the gel containing the target protein and transfer it to a membrane. Wash the membrane with TBST and block it for 1 hour with blocking solution. Incubate with diluted primary antibody overnight. Wash the membrane with TBST and incubate with secondary antibody at room temperature for 1 hour. Wash the membrane again with TBST, develop with ECL developer, acquire images, and measure HIF-1α levels.

[0135] The experimental results are as follows Figure 16 As shown: compared with the control group, the expression of HIF-1α protein in the model group was significantly increased; compared with the model group, the expression of HIF-1α protein in the drug-treated group was significantly decreased in a dose-dependent manner.

[0136] Example 17

[0137] Effects of 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol on α-smooth muscle actin (α-SMA), type I collagen (COL-Ⅰ), and E-cadherin in HK-2 cells induced by TGF-β1

[0138] Experimental method: HK-2 cells in the logarithmic growth phase were selected and digested with trypsin to prepare a concentration of 1.5×10 5 Cells were plated in a 12-well plate using a cell suspension of 100 μg / mL. Experimental, negative, and blank groups were assigned, and all plates were incubated overnight at 37°C in a 5% CO2 incubator. Experimental groups were stimulated with 10 ng / mL TGF-β1 for 24 hours. 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol was added to the plate using complete culture medium at concentrations of 10 μM, 20 μM, and 40 μM. An equal volume of complete culture medium was added to the control group, with triplicate wells per group treated for 24 hours. RNA was extracted using the SteadyPure Rapid RNA Extraction Kit. Total RNA extracted from each group was reverse transcribed using the EvoM-MLV Reverse Transcription Premix Kit. RT-qPCR was performed using the SYBR Green Pro Taq HS Premix qPCR Kit. After the RT-PCR reaction, the levels of α-SMA, COL-I, and E-cadherin were calculated using the 2-ΔΔCT method.

[0139] Table 1 Primer sequences for RT-PCR

[0140]

[0141] The experimental results are as follows Figure 17As shown in the results, compared with the control group, the mRNA expression levels of α-SMA and COL-Ⅰ were significantly increased (p<0.0001), and the mRNA expression level of E-cadherin was significantly decreased (p<0.001). After treatment with 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol, the mRNA expression levels of α-SMA and COL-Ⅰ in HK-2 cells were significantly decreased (p<0.0001), while the mRNA expression level of E-cadherin was significantly increased (p<0.0001~0.001). This indicates that 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol can effectively inhibit TGF-β1-induced cell transformation and collagen deposition, thereby alleviating the progression of renal fibrosis.

[0142] Example 18

[0143] Effects of 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol on phosphorylated phosphatidylinositol 3-kinase (p-PI3K) and phosphorylated protein kinase B (P-AKT) in HK-2 cells induced by TGF-β1

[0144] Experimental method: HK-2 cells in the logarithmic growth phase were selected and digested with trypsin to prepare a concentration of 3×10 5A cell suspension of 100 μg / mL was seeded in 6-well plates. Experimental, negative, and blank groups were set up, and all plates were incubated overnight at 37°C in a 5% CO2 incubator. Experimental groups were stimulated with 10 ng / mL TGF-β1 for 24 hours. 6-(3,4-dihydroxyphenyl)-2,3-naphthalenediol was prepared in complete culture medium at concentrations of 10 μM, 20 μM, and 40 μM. An equal volume of complete culture medium was added to the control group, and three wells per group were plated for 24 hours. The supernatant was discarded, and the culture medium was washed thoroughly with PBS. Pre-prepared cell lysis buffer was added, with 150 μL of lysis buffer added to each well. Mix thoroughly by pipetting, and then refrigerate at 4°C for half an hour to fully lyse the cells. After lysis, the cells were transferred to centrifuge tubes and centrifuged at 12,000 rpm for 10 minutes at 4°C. The supernatant was collected to obtain total protein, and 1 / 4 volume of loading buffer was added. The cells were boiled at 100°C for 5 minutes, then aliquoted and stored at -80°C for later use. Protein concentration was measured using a BCA kit. After preparing the electrophoresis tape, add the marker and sample for electrophoresis. After electrophoresis, cut the gel containing the target protein and transfer it to a membrane. After transfer, wash the membrane with TBST and block it with blocking buffer for 1 hour. Incubate with diluted primary antibody overnight. After washing with TBST, incubate with secondary antibody at room temperature for 1 hour. Wash the membrane again with TBST, develop with ECL developer, acquire images, and measure p-PI3K and p-AKT levels.

[0145] The experimental results are as follows Figure 18 As shown: Compared with the control group, the expression of p-PI3K (p<0.01) and P-AKT proteins in the model group increased; compared with the model group, the expression of p-PI3K and P-AKT proteins in the drug administration group was significantly decreased (p<0.001).

Claims

1. Use of a naphthalene diol derivative or a pharmaceutically acceptable salt thereof in the preparation of a drug for treating chronic kidney disease, wherein the naphthalene diol derivative is represented by formula (I): in, R1, R2, R 11 、R 12 Each is independently selected from hydrogen, C1-6 alkyl, C3-10 cycloalkyl, C1-6 acyl, phenyl or saccharyl; R3, R4, R5, R6, R7, R8, R9, R 10 Each is independently selected from hydrogen, C1-6 alkyl, C3-10 cycloalkyl, C1-6 acyl, phenyl, saccharide, halogen, amino, hydroxyl, aldehyde or carboxyl.

2. The use according to claim 1, characterized in that The naphthalene diol derivative is 6-(3,4-dihydroxyphenyl)-2,3-naphthalene diol (Formula II) 3. Use of a Dytiscidae extract in the preparation of a drug for treating chronic kidney disease, characterized in that The Dytiscidae extract comprises the naphthalene diol derivative according to claim 1 or 2.

4. The use according to any one of claims 1 to 3, characterized in that The chronic kidney disease includes chronic kidney disease induced by factors such as drugs, diabetes, hypertension, glomerulonephritis, etc.

5. The use according to claim 4, characterized in that The drugs include doxorubicin, which can induce chronic kidney disease.

6. The use according to any one of claims 1 to 3, characterized in that The chronic kidney disease includes chronic kidney disease caused by abnormal regulation of HK-2 in renal tubular epithelial cells induced by lipopolysaccharide (LPS), hydrogen peroxide (H2O2) or transforming growth factor-β1 (TGF-β1).

7. A drug for treating chronic kidney disease, characterized in that: The active ingredients of the medicine include Dytiscus spp. extract and / or naphthalene diol derivatives.

8. The drug for treating chronic kidney disease according to claim 7, characterized in that The concentration of the naphthalene diol derivative in an in vitro cell model is 1-50 μM, and the dosage of the Dytiscus spp. extract in an animal body is 50-300 mg / kg.

9. The drug for treating chronic kidney disease according to claim 8, characterized in that The concentration of the naphthalene diol derivative in an in vitro cell model is 2.5-40 μM, and the dosage of the Dytiscus spp. extract in an animal body is 55-220 mg / kg.

10. An inhibitor for a chronic kidney disease-related signaling pathway, characterized in that: The active ingredients of the inhibitor include a water slug extract and / or a naphthalene diol derivative, and the signal pathway includes a phosphatidylinositol 3-kinase (PI3K)-protein kinase B (Akt), a mitogen-activated protein kinase (MAPK), and a hypoxia-inducible factor-1 (HIF-1) signal pathway.

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